This project seeks to clarify molecular pathways that activate fetal hemoglobin or prevent its silencing as well as to define or develop novel transactivators of fetal hemoglobin. The net result of this study will be an increased understanding of globin regulation and novel molecular targets and therapeutic strategies to treat b-chain hemoglobinopathies. The ability to selectively manipulate the transcription of genes controlling the -globin locus is anticipated to have a significant impact on both our understanding of this locus as well as the treatment of diseases associated with mutations in this locus. The study proposed here capitalizes on our development of designed transcription factors that enable the transcription of endogenous genes to be either activated or repressed. Currently no other gene therapy strategy provides the means of effectively knocking out the expression of an endogenous gene- for example knocking out a silencer of g-globin expression. Polydactyl zinc finger proteins can now be prepared that recognize 18 bp DNA target sequences with high affinity and specificity. When fused to activation or repression domains, these proteins become potent regulators of the transcriptional activity of the target gene. This proposal focuses on the use of our transcriptional regulators to specifically modulate transcription of the b-globin locus with the aim of up-regulating fetal hemoglobin. We aim to explore the potential of targeted gene modulation as a gene-based therapeutic strategy for the treatment of b-hemoglobinopathies as well as a unique gene discovery tool for the identification of novel transcriptional modifiers of this locus and the validation of existing ones. A novel genome-wide transcriptional modulation strategy will be applied to the search for new targets for therapeutic intervention. With selective and potent transcriptional regulators we will address the therapeutic potential of controlling the globin locus in animal models. It is anticipated that the results of this work will provide a novel approach to study the molecular mechanisms of hemoglobinopathies as well as new strategies to treat them.